SUMOs are ubiquitin-related proteins that are posttranslationally and covalently conjugated to a large number of other cellular proteins, thereby regulating a variety of essential functions. In organisms ranging from yeast to man, SUMO modification regulates processes that include progression through the cell cycle, DNA synthesis and repair, chromosome segregation, initiation of transcription and nucleocytoplasmic transport. Although tremendous progress has been made over the past 5 years in understanding how SUMO is attached to substrates and in identifying cellular processes regulated by SUMO modification, many fundamental questions remain unanswered. Relatively little is known about how specific proteins are recognized and modified in response to the changes in cell growth or developement, or what specific signals facilitate this regulation. Also, the specific effects that SUMO modification has on substrates, and how these effects translate into changes in cell function, are poorly understood. The objectives of this proposal are to develop a more complete understanding of the cellular functions of SUMO modification and to understand how these functions are regulated and manifested at the molecular level. In this renewal application, the proposed experiments focus on: (1) defining the functions of SUMO-2/3 modification during mitosis (2) defining how SUMO-2/3 modification is regulated relative to SUMO-1 and to stages of the cell cycle (3) defining how SUMO-1 and SUMO-2/3 modification affects protein function. Importantly, all of the cellular processes regulated by SUMO conjugation are essential for normal cell growth and differentiation. Consequently, components of the SUMO conjugation pathway are associated with a variety of human diseases. SUMO conjugation plays a role in the processing of the amyloid precursor protein that is central to Alzheimer's disease, and it contributes to the pathology of Huntington's disease through modification of the Huntington protein. Due to functions in DNA repair and cell cycle control, the SUMO conjugation pathway may also contribute to the development of a variety of human cancers. Links to SUMO conjugation and susceptibility to type I diabetes have also been reported. Understanding how SUMO conjugation affects protein function at the moelcular level, and how it regulates cellular processes, is essential to defining how it roles in human diseases.